Figure 2 | Scientific Reports

Figure 2

From: Mechanisms that allow vaccination against an oncolytic vesicular stomatitis virus-encoded transgene to enhance safety without abrogating oncolysis

Figure 2The alternative text for this image may have been generated using AI.

An oncolytic vesicular stomatitis virus-vectored booster vaccine induced acute lymphopenia, which was evident in the transgene-specific T cell subset. C57BL/6 mice with orthotopic B16-F10 melanomas (2.5 × 105 cells injected intradermally) were unvaccinated, vaccinated intramuscularly with 1 × 108 IU of an adenovirus encoding human dopachrome tautomerase (Ad-hDCT) or primed with Ad-hDCT four days post-challenge, followed ten days later by intravenous administration of 1 × 109 pfu of vesicular stomatitis virus expressing hDCT (VSVΔm51-hDCT). Six- and twenty-four-hours post-VSV-hDCT, blood and saline-perfused spleens and tumors were harvested to quantify (a) lymphocytes (CD45+ cells in the lymphocyte gate shown in supplementary Fig. 1) by flow cytometry. (b) Similarly, hDCT-specific CD8+ T cells (defined as shown in supplementary Fig. 1) were quantified six hours post-VSV-hDCT. (c) The lymphopenic effect of oncolytic virotherapy was most prominent in the CD3loCD8lo blood-derived T cells (arrows), as demonstrated in the representative dot plots. (d) The ratios of CD3loCD8lo:CD3hiCD8hi T cells are shown. (e) Representative dot plots for four Ad-hDCT-vaccinated mice (10 days post-immunization) are shown after flow cytometric analysis following re-stimulation of blood-derived leukocytes with the DCT180–188 peptide. Interferon-gamma-producing T cells are overlaid in blue on the plots that show expression of CD3 and CD8. This is to demonstrate that activated antigen-specific T cells almost exclusively have a CD3loCD8lo phenotype. (f) The percentage of total CD8+ T cells that were specific for the immunodominant epitope from the adenoviral backbone (FAL) versus the melanoma-associated antigen (SVY) in blood. Controls were unvaccinated. These data were generated using an intracellular cytokine staining assay after re-stimulation with the peptides, followed by flow cytometric analysis. All graphs depict means and standard errors, with data pooled from three experiments (n = 16 unvaccinated control mice and n = 10 mice for both vaccinated groups). Results were analyzed by one-way analysis of variance with Tukey’s multiple comparison tests.

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